Therapeutic preparation of iron

ABSTRACT

4. A THERAPEUTIC COMPOSITION FOR TREATING IRON DEFICIENCY ANEMIA COMPRISING A NON-TOXIC, STERILE, COLLODIAL AQUEOUS SOLUTION OF A SUBSTANTIALLY NON-IONIC COMPLEX OF FERRIC HYDROXIDE WITH A DEXTRAN HAVING AN AVERAGE INTRINSIC VISCOSITY AT 25*C. OF ABOUT 0.025 TO ABOUT 0.25, SAID SOLUTION HAVING A PH OF ABOUT 4 TO ABOUT 8.5.

United States Patent Ofiice Re. 27,240 Reissued Nov. 23, 1971 27,240THERAPEUTIC PREPARATION OF IRON Eric London, Fareham, Hampshire, andGeorge Daniel Twigg, Goostrey, Cheshire, England, by FisonsPharmaceuticals Limited, Loughborough, Leicestershire, England,assignee, assignors to Fisons Limited, Felixstowe, Suffolk, England NoDrawing. Original No. 2,820,740, dated Jan. 21, 1958, Ser. No. 412,405,Feb. 24, 1954. Reissue No. 24,642, dated Apr. 28, 1959, Ser. No.765,693, Sept. 22, 1958. This application for reissue Oct. 7, 1969, Ser.No. 871,812 Claims priority, application Great Britain, Feb. 27, 1953,Int. Cl. A61]: 27/00 US. Cl. 424180 15 Claims Matter enclosed in heavybrackets [II appears in the first reissue but forms no part of thisreissue specification; matter printed in italics indicates the additionsmade by the first reissue; matter printed in bold face (except referencefigures and claim numbers) indicates the additions made by this reissueThis invention relates to the manufacture of colloidal therapeuticpreparations containing iron suitable for use in parenteral injectionand of the type called generically colloidal injectable ironpreparation.

It is well-known that the so-called saccharated oxide of ironpreparations can be used for injection purposes in the treatment ofiron-deficiency anemia. These preparations consist essentially ofaqueous solutions containing ferric hydroxide in colloidal form andstabilised by the presence of sugars (especially sucrose), of sugarmixtures, optionally modified by a preliminary heat-treatment, thecomponents being admixed in the presence of a suitable alkali.

While such materials may be injected intravenously without toxiceffects, they are found to be disadvantageous for intramuscular orsubcutaneous injection, since, owing to the high osmotic pressure andthe possible alkalinity of this type of solution, intramuscularinjection usually results in considerable pain and inflammation at thesite of the injection, even when sub-therapeutic doses are administered.The unsnitability of such products for intramuscular injection isrendered even more obvious by the fact that they are not absorbedappreciably when injected by this route.

It is considered desirable for an iron solution intended forintramuscular injection to be able to satisfy the followingrequirements:

(a) No effect of the pH of the body fluids;

(b) Isotonicity with the tissue fluid;

() Stability in presence of protein and electrolytes;

(d) Ready availability for haemoglobin synthesis;

(e) Rapid absorption coupled with a low rate of excretion;

(f) Maximal iron content in minimal volume, i.e. 3 to 5% elemental ironin solution;

(g) low toxicity;

(h) Reproducibility;

(i) Stability on storage.

The principal object of our invention is to provide more particularlyfor the treatment of iron-deficiency anemia a colloidal ironpreparation, which is especially suitable for intramuscular injection(although adaptable also for intravenous use) and is well-toleratedwithout the onset of undesirable local or general side-efiects. Afurther object is to obtain an iron-containing colloid,

essentially free from iron ions, which can be administered in dosageadequate for producing the required therapeutic response and from whichthe iron is absorbed rapidly into the circulation with absence ofuntoward systemic reactions, e.g. vasodilation, headache, lumbar pain,vomiting and loss of consciousness.

With the foregoing objects in view, the present invention provides atherapeutically effective iron preparation consisting essentially of acolloidal substantially non-ionic ferric hydroxide-partiallydepolymerized dextran complex, believed to be novel in itself. Acolloidal iron preparation may be produced according to the invention byreacting, with such heating as may be necessary, a solution of apartially depolymerized dextran, a solution of suspension of a ferriccompound (which expression covers for example the ferric compoundshereinafter specified or a mixture of ferric compounds), together withalkali, to form a colloidal solution from which any undissolved mattermay be separated and whose pH is or, if necessary, is adjusted tobetween 4.0 and 11.0 within which range the preparation appears to becapable of retaining its stability in water for reasonable periods oftime. However, the pH of the preparation for injection may be, or ifnecessary, may be adjusted to between 5.5 and 8.5 and preferably 6.5.

In carrying out this method, the partially depolymerized dextran mayeither be dissolved in the solution or suspension of the ferric compoundwith subsequent addition of the alkali, or alternatively, the dextrancan be dissolved in the alkali, to which the ferric compound in solutionor suspension is then added.

Though the molecular structure of the product is not yet known withcertainty, and it can therefore be defined only in empirical terms, ourparenteral iron preparation may possibly consist of an aqueous solutionof colloidal ferric hydroxide complexed with enough of the partiallydepolymerized dextran to stabilise the solution satisfactorily for thepurpose of injection but not so much ferric hydroxide as to require anexcess of the dextran over the requirements of such injection in otherrespects, e.g. as regards the consistency of the solution. If the ironcontent is excessive, the amount of the dextran needed to stabilise itis so great that the resultant high viscosity of the product renders itunsuitable for use as a parenteral iron preparation and a minimumdextraniiron ratio exists below which the iron can no longer bestabilised so as to be suitable for an injection.

The partially-depolymerized dextran which we use as a starting-materialis derived from raw dextran obtained according to known methods bygrowing under carefullycontrolled conditions of temperature, appropriateorganisms, especially Leuconostoc mesenteroides, syn. Betacoccusarabinosaceous in a suitable nutrient medium containing a highproportion of sucrose. It is further known that the highly polymerisedraw dextran gives rise on partial degradation as, for example, bytreatment with dilute mineral acid, to simpler polymeric forms fromaqueous solutions of which (being polydisperse), fractions of differentaverage molecular weight can be precipitated by adding a suitablewater-miscible organic liquid, such as methyl alcohol, ethyl alcohol oracetone. These degradation products consist of polymerised glucoseresidues, joined predominantly by t1-1I6 and, to a lesser extent, by al:4 linkages.

A fraction of this kind is suitable as a starting-material for thepurpose of our invention, but the molecular range of the partiallydepolymerized dextran is selected from the standpoint of the physicalcharacteristics such as intrinsic viscosity, of the solution containingthe minimum concentratiin of the dextran needed to ensure continuedstability of the parenteral iron solution, rather than from thecustomary one of the physiological effects of the dextran itself, whichare here of secondary importance.

In the present context the intended meaning of intrinsic viscosity isthe limiting value of specific viscosity divided by concentration atinfinite dilution measured by the following method. The fiow time of atleast three solutions of different concentrations (all less than w./v.dextran) is determined in an Ostwald viscometer. The flow time of thesolution divided by the How time for water at the same temperature,gives the relative viscosity of the dextran solution at the givenconcentration. The specific viscosity of a dextran solution at a givenconcentration is obtained by subtracting 1.0 from the value for relativeviscosity. For each concentration of dextran solution, the factor(specific viscosity divided by concentration) is calculated and thisfactor is plotted against the percentage concentration of the solutionfrom which it was obtained. By extrapolating the graph obtained to zeroconcentration, the limiting value of the factor (specific viscositydivided by concentration) is obtained. This value is known as theintrinsic viscosity of the material in solution. Due to the fact thatthe fractions of dextran described are polydisperse, the term averageintrinsic viscosity is used.

To avoid any possible misinterpretation, the term average intrinsicviscosity refers to the dextran itself and not to an aqueous solution ofdextran, or an aqueous solution of the ferric hydroxide-dextran complex.

We have found that stable iron solutions can be prepared from fractionsof dextran of intrinsic viscosity ranging from 0.025 to 0.5; but thequantity of iron stabilised per unit weight of dextran decreases as theintrinsic viscosity of the dextran colloid increases. The iron contentof a solution considered to be adequate for parenteral injection is atleast 2% and our investigations have shown that, in order to obtain astable preparation which possesses clinical utility the dextran selectedas a starting-material must have an upper limit of intrinsic viscositywhich does not appreciably exceed 0.2 and which more particularly has anintrinsic viscosity in the range from about 0.025 to about 0.25 at 25 C.A therapeutically-useful preparation containing from 3 to 5% elementaliron (corresponding to from 6 to 10% ferric hydroxide) can be obtainedfrom a fractionated dextran with an intrinsic viscosity of 0.03 to 0.06,when the partially depolymerized dextran content is approximately 30 to50%.

The colloidal ferric hydroxide may be formed in presence of thepartially depolymerized dextran by heating a suitable water-solubleferric salt and the dextran together in aqueous solution with excess ofalkali.

Suitable ferric compounds comprise: (i) water soluble ferric salts suchas the chloride, nitrate, sulphate or acetate and double salts such asferric ammonium sulphate, or their obvious chemical equivalents; (ii)ferric oxy-salts prepared by dissolving ferric hydroxide in a solutionof a ferric salt; dialysed iron solution B.P.C. and freshlyprecipitatedWashed ferric hydroxide; (iii) any ferric compound which, when renderedalkaline in the presence of dextran, gives rise to ferric hydroxide.

The alkali of choice for the purpose of this invention is sodiumhydroxide, but other suitable alkalis comprise the hydroxides oflithium, potassium and ammonium; the carbonates of lithium, sodium andpotassium and their obvious chemical equivalents.

Stable preparations appropriate for intramuscular injection can also beobtained according to our invention from ferric citrate, ferric ammoniumcitrate and ferric glycerophosphate, but in these instances we haveshown it to be desirable definitely to use a caustic alkali, forexample, sodium hydroxide, as the agent for rendering the mixturealkaline during the preparation thereof.

We have further found that our novel colloidal injectable ironpreparations can be purified from the electrolytes which are formedsimultaneously as by-products by utilising either of the followingtechniques:

(i) Subjecting a preparation according to the invention to dialysisagainst running water until the desired osmotic pressure has beenobtained;

(ii) Mixing a preparation according to the invention with a suitablewater-miscible solvent such as methyl alcohol, ethyl alcohol or acetonein quantity sufiicient to separate the colloid, separating the latterfrom solution and re-dissolving it in distilled water to the requiredconcentration.

It sometimes happens that a parenteral iron preparation obtained by theimproved process is slightly hypotonic; thus the freezing-pointdepression of a colloidal solution containing about 5% elemental iron,as normally prepared for intramuscular injection, is frequently lessthan 0.53 C. (the approximate depression obtained with isotonic saline):in such a case, a suitable amount of an appropriate substance such assodium chloric or glucose may be added to raise the freezing-pointdepression value of the preparation to 0.53 C. and thereby render itsubstantially isotonic with blood.

If the pH of a colloidal iron preparation in accordance with ourinvention falls outside the range 5.5 to 8.5, there can be added, priorto sterilization, sutficient acid or alkali, as the case may be, toensure that a pH value falling within these limits, and preferably 6.5,is finally attained for clinical use.

Sterilization of the preparation made in accordance with the presentinvention can be effected by autoclaving the said preparations in theirfinal containers, as, for example, ampoules, for 30 minutes at a steampressure of 10 lbs. per square inch, corresponding to a temperature ofC.

When colloidal solutions prepared in accordance with our invention aredehydrated as, for example, by evaporation under reduced pressure or,alternatively, by the separation and subsequent desiccation of theproduct obtained on precipitating the colloid by adding a suitablewatermiscible solvent such as methyl alcohol, ethyl alcohol or acetone,there can be obtained solidified preparations suitable for the immediatereconstitution of colloidal injectable iron solutions by the addition ofdistilled water.

Tested for intravenous toxicity in mice, a preparation obtainedaccording to our investigation may show an LD value in excess of 600mg./kilo: this result compares most favourably with a commercialsaccharated oxide of iron preparation which, when tested under similarconditions, indicated an LD of 300 mg./ kilo.

On injecting a colloidal parenteral iron preparation made in accordancewith this invention into mice by the intramuscular route it has not beenfound possible to administer a dosage sufficiently large to kill any ofthe test animals (mice or rats) using the highest practicalintramuscular dosage of the material, viz: 5 mls./kilo (corresponding to250 mgms. Fe/ kilo).

The following are examples by way of illustration only of preparationsin detail according to the invention:

EXAMPLE 1 To 25 g. partially-depolymerized dextran (intrinsic viscosity:0.07), dissolved in water (50 ml.), was added sodium hydroxide (15 g.)in water (25 ml.), followed by 40 ml. of 30% w./v. aqueous ferricchloride. The mixture was heated to boiling for about 15 minutes andallowed to cool to room temperature. The solid that remained undissolvedwas then removed by centrifugation and the solution dialysed againstrunning water for approximately 24 hrs, using tubing made of materialknown under the registered trademark cellophane. The dialysed solutionwas concentrated under reduced pressure to yield a clear, stablesolution containing the equivalent of 4.15% elemental iron. The productwas again filtered and sterilised by autoclaving at a steam pressure oflbs. per square inch for 30 minutes, its final pH value being 6.8. Thepreparation was found to have a freezing-point depression of 0.18" C.,compared with pure water, which on addition of 0.6% w./v aqueous sodiumchloride, was raised to 0.60 C.

This material when tested for intravenous toxicity in mice gave an LDvalue of approimately 800 mg./kilo.

EXAMPLE 2 To 25 g. partially-depolyrnerised dextran (intrinsicviscosity: 0.05) dissolved in water (50 ml.) was added so dium hydroxideg.) in water (25 ml.), followed by ferric citrate (22 g.) in water (200ml.). The mixture was heated for approximately 2 hrs. at 65 C., withstirring and the resultant dark red solution was filtered and cooled.The filtrate was stirred with 95% ethyl alcohol and the treaclyprecipitate separated, prior to its being redissolved in about 1 litredistilled water. From this aqueous solution, the material was againprecipitated under the same conditions, to be once more dissolved indistilled water. The filtered aqueous solution was evaporated underreduced pressure at 45 C., until the solution contained 5% Fe, asdetermined by assay.

EX A MPLE 3 400 g. of dextran having an intrinsic viscosity of 0.04, and103 g. of anhydrous sodium carbonate are dissolved in 800 ml. of hotwater. This solution is cooled and to it 400 ml. of a solution of ferricchloride (containing 100 g. 'FeCl is added with stirring. After theliberated carbon dioxide has escaped, the resulting solution containingcolloidal ferric hydroxide stabilized with dextran and also sodiumchloride produced during the reaction, is dialysed against water, using,for example, cellophane as a membrane. When its sodium chlorideconcentration has in this way been reduced to about 0.2%, the solutionis concentrated under atmospheric pressure until the volume is reducedto about 600 ml. The preparation is then adjusted to a 5% content ofiron and sterilized by autoclaving in the final containers.

We claim:

1. A composition comprising a substantially non-ionic complex of ferrichydroxide with a dextran having an average intrinsic viscosity at 25 C.of about 0.025 to about 0.25, said complex being stable in contact withWater.

2. A therapeutic composition comprising a stable aqueous solution of asubstantially non-ionic complex of ferric hydroxide with a dextranhaving an average intrinsic viscosity at 25 C. of about 0.025 to about0.25.

3. A composition comprising a substantially non-ionic complex of ferrichydroxide with a dextran having an average intrinsic viscosity at 25 C.of about 0.03 to 0.06, said complex being stable in contact with water.

4. A therapeutic composition for treating iron deficiency anemiacomprising a non-toxic, sterile, colloidal aqueous solution of asubstantially non-ionic complex of ferric hydroxide with a dextranhaving an average intrinsic viscosity at 25 C. of about 0.025 to about0.25, said solution having a pH of about 4 to about 8.5.

5. A composition according to claim 4 having at least 2% elemental iron.

[6. The process which comprises parenterally administering to an animala therapeutic amount of a substantially non-ionic complex of ferrichydroxide with a dextran having an average intrinsic viscosity at 25 C.of about 0.025 to about 0.25.]

[7. The process of claim 6 in which the parenteral administration isintramuscular] [8. The process of preparing a substantially non-ioniccolloidal ferric hydroxide-dextran complex which comprises combining, incontact with water, a dextran having an average intrinsic viscosity at25 C. of about 0.025 to about 0.25 with ferric hydroxide, said ferrichydroxide being formed in situ in contact with the dextran by a doubledecomposition reaction between an ionizable ferric salt and an alkalibase] [9. The process of claim 8 in which the resulting ferrichydroxide-dextran complex in water is dialyzed to remove electrolytes][10. The process of claim 8 in which the pH is adjusted to the rangefrom about 4 to about 8.5.]

[11. The process of claim 8 in which the resulting ferrichydroxide-dextran complex is purified by precipitation with a watermiscible solvent, the precipitate is separated, and redissolved inwater] 12. A composition comprising a substantially nonionic complex offerric hydroxide with a dextran having an average intrinsic viscosity at25 C. of about 0.03 to 0.06, said complex containing at least 2%elemental iron, and being stable in contact with water.

13. A therapeutic composition in a final sterilized container fortreating iron deficiency anemia, said therapeutic composition comprisinga substantially non-ionic complex of ferric hydroxide with a dextranhaving an average intrinsic viscosity at 25 C. of about 0.025 to 0.25,said complex being stable in contact with water.

14. An ampoule containing a therapeutic composition for treating irondeficiency anemia, said therapeutic composition comprising a non-toxic,sterile, colloidal aqueous solution of a substantially non-ionic complexof ferric hydroxide with a dextran having an average intrinsic viscosityat 25 C. of about 0.025 to about 0.25, said solution having a pH ofabout 4 to about 8.5.

15. The process which comprises intramuscularly administering to ananimal a therapeutic amount of a substantially non-ionic complex offerric hydroxide with a dextran having an average intrinsic viscosity at25 C. of about 0.025 to about 0.25.

16. A therapeutic composition comprising a substantially non-ioniccomplex of ferric hydroxide with a dextran in a final sterilizedcontainer for treating iron deficiency anemia in animals, said dextranhaving an average intrinsic viscosity at 25 C. of about 0.025 to 0.25,said complex being stable in contact with water.

17. A therapeutic composition comprising a substantially non-ioniccomplex of ferric hydroxide with a dextran in an ampoule for treatingiron deficiency anemia in animals, said dextran having an averageintrinsic viscosity at 25 C. of about 0.025 to 0.25, said complex beingstable in contact with water.

J 8. A therapeutic composition comprising a substantially non-ioniccomplex of ferric hydroxide with a dcxtran having an average intrinsicviscosity at 25 C. of about 0.025 to 0.25, said complex being stable incontact with water and having clinical use for treating iron deficiencyanemia with the absence of untoward systemic reactions, such asheadaches, and lumbar pain.

19. A therapeutic composition comprising a substantially non-ioniccomplex of ferric hydroxide with a dextran in a final sterilizedcontainer, said dextran having an average intrinsic viscosity at 25 C.of about 0.025 to 0.25, said complex being stable in contact with waterand having clinical use for treating iron deficiency anemia with theabsence of untoward systemic reactions, such as headaches, and lumbarpain.

20. A therapeutic composition comprising a substantially non-ioniccomplex of ferric hydroxide with a dextran in an ampoule, said dextranhaving an average intrinsic viscosity at 25 C. of about 0.025 to 0.25,said complex being stable in contact with water and having clinical usefor treating iron deficiency anemia with the absence of untowardsystemic reactions, such as headaches, and lumbar pain.

21. A therapeutic composition for treating iron deficiency anemia,comprising a substantially non-ionic complex of ferric hydroxide with adextran having an average 7 8 intrinsic viscosity at 25 C. of about 0.03to 0.06, said OTHER REFERENCES complex being stable in contact withwater and contain- Zief et a1 LA'C Apr 20 1952 pp 2126 7 ing about 5%elemental iron. I l

Manufacturing Chemist, vol. 23:2 February 1952, pp.

References Cited 5 Physician's Bulletin, May 1952 pp. 4144.

The following references, cited by the Examiner, are

of record in the patented file of this patent or the original SAM ROSEN,Primary Examiner patent.

UNITED STATES PATENTS 2,518,135 8/1950 Gaver. 421F147 2,572,923 10/1951Gaver.

2,609,368 9/1952 Gavel.

12;;3? UNITED STATES PATENT OFFICE CERTEFICATE 0F CORRECTION ReissuePatent No. Re Z7 Z40 Dated November 23 1971 Inventor(s) Eric London andGeorge Daniel Twigg It: is certified that error appears in theabove-identified patent and that said Letters Patent are herebycorrected as shown below:

I- Y Column 3, line 4 "concentratiin" should be concentration- Claims 13through 21 should be printed in bold face to indicate the additions madeby this reissue Signed and sealed this 2nd day of May 1 972.

(SEAL) e Attest;

EDWARD M.F'LETC HER,-JR. ROBERT GOTTSCHALK attesting OfficerCommissioner of Patents Page 1 of l

4. A THERAPEUTIC COMPOSITION FOR TREATING IRON DEFICIENCY ANEMIACOMPRISING A NON-TOXIC, STERILE, COLLODIAL AQUEOUS SOLUTION OF ASUBSTANTIALLY NON-IONIC COMPLEX OF FERRIC HYDROXIDE WITH A DEXTRANHAVING AN AVERAGE INTRINSIC VISCOSITY AT 25*C. OF ABOUT 0.025 TO ABOUT0.25, SAID SOLUTION HAVING A PH OF ABOUT 4 TO ABOUT 8.5.